Note : Les descriptions sont présentées dans la langue officielle dans laquelle elles ont été soumises.
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UTILIZATION OF VINPOCETINE TO AVOID COMPLICATIONS IN
PARTICULAR THOSE ASSOCIATED TO HEARING WHICH OCCUR WITH
EPILEPSY, AND TREATMENT THEREOF
TECHNICAL FIELD
The present invention is related with the use of vinpocetine and the
derivates developed from its formula that maintain the same effects for the
treatment of epilepsy and its complications, particularly those related with
the
auditory pathway.
BACKGROUND
One major problem of epilepsy is the deleterious cognitive and
behavioural consequences caused by the illness (Prevey et al. 1998 Epilepsy
Res. 30: 1; Jokeit and Ebner 1999 J. Neurol. Neurosurg. Psychiatry 67: 44;
Theodore et al. 1999 Neurology 52: 132; Meador 2001 Epilepsy Behav. 2:
307) as well as by its treatment with the available antiepileptic drugs (Gates
2000 Epilepsy Behav. 1: 153; Kwan and Brodie 2001 Lancet 357: 216;
Brunbech and Sabers 2002 Drugs 62: 593; Schmidt 2002 Epilepsy Res. 50:
21).
The implication of the auditory brainstem nuclei in the pathophysiology
of generalized epilepsy is indicated by the alterations in the latencies
and/or
amplitudes of the later waves of the auditory brainstem response (ABR)
observed in patients with generalized epilepsy (Rodin et al. 1982 Clin.
Electroencephalogr. 13: 154; Mervaala et al. 1986 Epilepsia 27: 542; Phillips
et al. 1990 Clin. Electroencephalogr. 21: 135; Soliman et al. 1993 Ear Hear
14: 235; Kohsaka et al. 1999 Brain Res. 837: 277; Kohsaka et al. 2001 Brain
Res. 903: 53). The acute epilepsy induced by two convulsive agents in non
medicated animals is accompanied of abnormalities in the later waves of the
ABR and by a marked hearing decline (Nekrassov and Sitges 2003 Epilepsy
Res. 53: 245).
ABRs are far field-evoked potentials that consist of several waves that
occur within 10 ms after the auditory stimulus. Since changes in the later
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waves of the ABR indicate alterations of specific nuclei of the auditory
brainstem (Hughes, J.R., Fino, J.J., 1985 J. Clin. Neurophysiol. 2: 355), ABRs
are commonly used in the clinical diagnosis of retro-cochlear lesions. In
addition, the ABR threshold is used in the clinical diagnosis of the hearing
sensitivity, because stimuli of progressively higher intensity (in dB) are
needed for evoking the ABR while the hearing sensitivity declines. Therefore
elevations in the ABR thresholds are an objective determination of hearing
deterioration.
Antiepileptic drugs, including carbamazepine, valproate, phenytoin,
phenobarbital, clonazepam and vigabatrin, also cause abnormalities on the
waves of the ABR as well as hearing deficits (Mervaala et al. 1987
Electroencephalogr. Clin. Neurophysiol. 68: 475; Armon et al. 1990 Neurology
40: 1896; Hirose et al. 1990 Electroencephalogr. Clin. Neurophysiol. 75: 543;
Yuksel et al. 1995 Childs Nerv. Syst. 11: 474; De la Cruz and Bance 1999
Arch Otolaryngol Head Neck Surg. 125: 225; Zgorzalewicz and Galas-
Zgorzalewicz 2000 Clin. Neurophysiol. 111:2150).
Vinpocetine (ethyl apovincamine-22-oate) discovered during the late
1960s has successfully been used in the treatment of central nervous system
disorders of cerebrovascular origin for decades. In animal models of hypoxia
and ischemia vinpocetine exerts beneficial effects against neuronal damage
(King 1987 Arch. Int. Pharmacodyn. Ther. 286:299; Araki et al. 1990
Res.. Exp.. Med. 190:19).
More recently vinpocetine has also been used for ameliorating memory
on the basis of previous studies in animals and humans (Subhan and
Hindmarch 1985 Eur. J. Clin. Pharmacol. 28: 567; Bhatti and Hindmarch 1987
Int. Clin. Psychopharmacol. 2: 325; DeNoble 1987 Pharmacol. Biochem.
Behav. 26: 183).
Vinpocetine is a Na+ channel blocker (Erdo et al. 1996 Europ. J.
Pharmacol. 314:69). In brain isolated nerve endings we have shown that
vinpocetine selectively inhibits neurotransmitter release induced by an
increase in presynaptic Na+ channels permeability (Sitges and Nekrassov,
1999 Neurochem. Res. 24:1587; Trejo et al. 2001 Brain Res. 909:59). In
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addition we have shown that in the guinea pig in vivo vinpocetine exerts a
long term inhibition of the alterations in the ABR waves, the hearing loss and
the mortality induced by amikacin (Nekrassov and Sitges 2000 Brain Res.
868: 222) and other aminoglycoside antibiotics (unpublished results).
There is an uncovered medical need for the treatment of epilepsies.
Medication with antiepileptic drugs of either the "old and new generations"
although exerts a positive effect on seizure control (at least in about 70% of
epileptic patients), deteriorates the cognitive functions (Vermeulen and
Aldenkamp 1995 Epilepsy Res. 22: 65; Gates 2000 Epilepsy Behav. 1: 153;
Brunbech and Sabers 2002 Drugs 62: 593; Schmidt 2002 Epilepsy Res. 50:
21), that may aggravate the cognition decline caused by the illness (Prevey et
al. 1998 Epilepsy Res. 30: 1; Jokeit and Ebner 1999 J. Neurol. Neurosurg.
Psychiatry 67: 44; Theodore et al. 1999 Neurology 52: 132; Meador 2001
Epilepsy Behav. 2: 307). The antiepileptic drugs also cause alterations in the
waves of the ABR (Mervaala et al. 1987 Electroencephalogr. Clin.
Neurophysiol. 68: 475; Armon et al. 1990 Neurology 40: 1896; Hirose et al.
1990 Electroencephalogr. Clin. Neurophysiol. 75: 543; Yuksel et al. 1995
Childs Nerv. Syst. 11: 474; De la Cruz and Bance 1999 Arch Otolaryngol
Head Neck Surg. 125: 225; Zgorzalewicz and Galas-Zgorzalewicz 2000 Clin.
Neurophysiol. 111:2150), that can result in a hearing decline (Nekrassov and
Sitges 2003 Epilepsy Res. 53: 245); and fail to produce a perceptible impact
on the prophylaxis of the illness after the first seizure (Hernandez 1997
Trends Pharmacol. Sci. 18: 59; Temkin et al. 2001 Drugs 61: 1045; Schmidt
2002 Epilepsy Res. 50: 21).
The available antiepileptic drugs provoke several adverse secondary
effects that in many occasions lead the patients to stop the treatment,
because of the important alterations that limit their everyday life.
The present invention refers to the use of vinpocetine and the derivates
developed from its formula that maintain the same effects for preparing a drug
of choice in the treatment of epilepsies. The present invention describes the
beneficial action of vinpocetine for preventing the epileptic cortical
activity for
the ictal and post-ictal periods and for preventing the most important
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disturbances caused by the illness and aggravated by the available
antiepileptic drugs.
According to an aspect of the present invention, there is provided the
use of at least one of vinpocetine and derivatives thereof in the manufacture
of a medicament for antagonizing alterations in auditory brainstem response
(ABR) waves and hearing loss that accompanies epilepsy.
According to another aspect of the present invention, there is provided
use of at least one of vinpocetine and derivatives thereof for antagonizing
alterations in auditory brainstem response (ABR) waves and hearing loss that
accompanies epilepsy.
In accordance with another aspect, there is provided use of vinpocetine
for fabrication of a medicine useful in treatment of hearing loss and
prevention of the characteristic epileptic cortical activity for the ictal and
post-ictal periods accompanying epilepsy.
BRIEF DESCRIPTION OF THE FIGURES
Figure 1. This figure shows the ABR recordings taken in an
representative animal before (a) and 50 min after the injection of
pentylenetetrazole (PTZ) in an animal pre-injected with vehicle 4 hours before
PTZ (b), and in an animal pre-injected with vinpocetine 4 hours before PTZ
(c). The animal received a pure tone monaural stimulus of high frequency (8
kHz) and high intensity (100 dB) indicated by the arrow.
Figure 2. The figure shows that vinpocetine inhibits the increase in the
peak latencies of P2, P3 and P4 waves of the ABR induced by PTZ. The
latencies of the waves induced by stimuli of 100 dB at tone frequencies of 8
kHz (left graphs) and 4 kHz (right graphs) were determined before (Bef.) and
10, 20, 30 and 50 min after the injection of PTZ in control animals pre-
injected
with vehicle (black circles) and in animals pre-injected with 2mg/kg
vinpocetine (empty circles). Results are the mean SEM values of 8
independent animals.
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Figure 3. The figure shows that vinpocetine inhibits the changes on the
electroencephalogram (EEG) induced by PTZ for the ictal period. The EEG
recordings shown were taken in a representative animal before (top trace)
and about 2 min after the injection of PTZ in the animal pre-injected with
vehicle 4 hours before PTZ (middle trace) or pre-injected with vinpocetine 4
hours before PTZ (bottom trace).
Figure 4. The figure shows that vinpocetine inhibits the changes on the
EEG induced by PTZ for the post-ictal period. EEG recordings were taken in a
representative animal pre-injected with vehicle before (top trace) and 10, 20,
30 and 50 min after the injection of PTZ. EEG recordings were taken in a
representative animal pre-injected with vinpocetine before (top trace) and 10,
20, 30 and 50 min after the injection of PTZ.
Figure 5. The figure shows that vinpocetine inhibits the alterations in
the amplitude of the later ABR waves induced by another convulsive agent, 4-
4a
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aminopyridine (4-AP). The ABRs were induced by a stimulus of 100 dB at 8
kHz. It is shown that the progressive increase in the amplitude of the P3 wave
of the ABR observed after the injection of 4-AP in control animals (a) is
eliminated in the animals pre-injected with vinpocetine (b), and that the
marked decrease in the amplitude of the P4 wave of the ABR observed after
the injection of 4-AP in control animals (c) is considerably reduced in the
animals that received vinpocetine (d).
Figure 6. This figure shows that vinpocetine inhibits the changes on the
EEG induced by 4-AP for the ictal period. The EEG recordings shown in (a)
were taken in a representative animal before and about 20 min after the
injection of 4-AP in an animal pre-injected with vehicle. The EEG recordings
shown in (b) were taken in a representative animal before and about 20 min
after the injection of 4-AP in the animal pre-injected with 2mg/kg
vinpocetine.
Figure 7. This figure shows that vinpocetine inhibits the changes on the
EEG induced by 4-AP for post-ictal period. The EEG recordings shown in (a)
were taken in a representative animal pre-injected with vehicle before (top
trace) and 30, 60 and 80 min after the injection of 4-AP. The EEG recordings
shown in (b) were taken in a representative animal pre-injected with
vinpocetine before (top trace) and 30, 60 and 80 min after the injection of 4-
AP.
DETAILED DESCRIPTION OF THE INVENTION
In a previous study we have shown that the alterations in the activity of
the lateral and the medial nuclei of the superior olivary complex reflected in
abnormalities in the parameters (amplitude and latency) of the later waves of
the ABR are connected with the hearing decline caused by epilepsy
(Nekrassov and Sitges 2003 Epilepsy Res. 53: 245).
The present invention demonstrates that vinpocetine inhibits the
alterations in the amplitude and latency of the later ABR waves, as well as
the
hearing decline and the characteristic epileptic cortical activity observed in
two
models of experimental animal epilepsy in vivo.
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Epilepsy can be induced in experimental animals in vivo either by
decreasing the cerebral inhibitory transmission or by increasing the cerebral
excitatory transmission. This can be achieved the injection of the GABA
antagonist, PTZ or the glutamate releaser, 4-aminopiridine (4-AP),
respectively. Our results in the guinea pig show that when vinpocetine is
administered at a concentration of 2 mg/kg i.p. 4 hours before the injection
of
PTZ or 1 hour before the injection of 4-AP, none of these convulsing agents is
capable to induce the alterations in amplitude and latency of the later ABR
waves, to provoke the hearing decline or to induce the epileptic cortical
activity.
In the guinea pig the P3 and P4 waves of the ABR express the activity
of the medial and the lateral superior olivary nuclei, respectively (Wada and
Starr 1983 Electroencephalogr. Clin. Neurophysiol. 56: 326; 56: 340; 56:
352). Among the nuclei of the superior olive, the P4 generator is determinant
in sound source localization (Tollin 2003 Neuroscientist 9: 127). Changes in
these late waves of the ABR indicate retro-cochlear alterations. Vinpocetine
cancels all the retro-cochlear abnormalities induced by PTZ or by 4-AP
(evidenced by the changes in P3 and/or P4 parameters) and by this mean
prevents the hearing decline induced by both convulsing agents.
The available antiepileptic drugs induce alterations in the parameters of
the ABR waves and hearing deficits that may aggravate the abnormalities and
the hearing loss induced by the illness. Medication with antiepileptic drugs
although exerts a positive effect on seizure control also causes secondary
adverse effects, among which cognition decline and hearing loss are
particularly important. Vinpocetine is Well tolerated and without
contraindications in humans at doses as high as 60 mg/day (Hindmarch et al.
1991 Int. Clin. Psychopharmacol. 6: 31). Our results indicate that vinpocetine
at a reasonable dose (2 mg/kg) completely cancels the ictal and post-ictal
cortical activity induced by PTZ and 4-AP, as well as the hearing loss induced
by the two convulsing agents. The contribution of hearing loss to the decline
exerted by epilepsy and by the classic antiepileptic drugs on cognition is
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prevented by vinpocetine, indicating the advantage that represents the use of
vinpocetine as antiepileptic over the classical antiepileptic treatments.
Another problem of the available antiepileptic drugs is that they fail to
produce a perceptible impact on epileptogenesis or the progression of the
illness. In a previous study we have shown that vinpocetine exerts a long term
(more than half a year) protective action over the changes in the ABR waves,
the hearing loss, and the mortality induced by the treatment with a high dose
of amikacin (Nekrassov and Sitges 2000 Brain Res. 868: 222), suggesting
that vinpocetine may also exert an important prophylactic action in the
treatment of epilepsy.
In summary, our findings indicate that vinpocetine prevents the
epileptic cortical activity for the ictal and post-ictal periods, as well as
the
alterations in the ABR waves, that result in hearing loss and contribute to
the
adverse effects caused by epilepsy and the available antiepileptic drugs on
cognition. These findings, along with the vinpocetine long term protective
action indicate that vinpocetine is a better alternative in the treatment and
prophylaxis of epilepsies.
EXAMPLES
Experiments were performed in pigmented adult male guinea pigs
initially weighing 349 38 g. ABRs recordings were used to evaluate the
hearing status of each animal and EEG recordings to evaluate changes in
cortical excitability. The ABR and EEG recordings were obtained following a
method that we have previously reported (Nekrassov and Sitges, 2003). The
Institutional Animal Use and Care Committee approved all experimental
procedures.
Three types of recordings were performed in the anaesthetized
animals, namely the ABR recordings elicited by a stimulus of high intensity
(100 dB), the ABR recordings for determination of the auditory threshold and
the EEG recordings.
Determination of ABR wave parameters. The latency and amplitude of
each wave component of the ABR elicited by a stimulus of 100 dB with pure
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tone frequencies of 4 and 8 kHz was measured in all the ABR recordings
obtained under the different experimental conditions at the specified times.
The latency of each ABR wave (in ms) refers to the time interval between the
onset of the auditory stimulus and the positive peak of the wave. The onset of
the stimulus is indicated by the vertical arrow at the bottom of the
recordings
on Fig. 1. The peak amplitude (in pV) of each wave of the ABR is the
difference between the positive peak of the wave and the reference baseline
(trace between the stimulus and the appearance of the first ABR wave on Fig.
1).
Determination of ABR thresholds. ABR recordings elicited by stimuli of
progressively lower intensity (in dB) were used for determining the hearing
threshold. Threshold is defined as the lowest stimulus intensity (in dB) at
which the P3 wave of the ABR could still be recorded in three consecutive
trials.
Student's t-test was used for the evaluation of the differences between
results obtained before and at the specified times after the injection of the
convulsing agents. The criterion for statistical significance for all measures
was P _< 0.05. All data are expressed as means standard error of the mean.
In figures and tables the symbol * is used for indicating statistic
significant
differences.
Example 1. Experimental design used for testing the effect of vinpocetine on
the changes on the ABR and EEG recordings induced by epilepsy resulting
from a reduced cerebral inhibitory transmission.
Eight male guinea pigs were entered into this study. PTZ was dissolved
in saline and vinpocetine in a saline acidified (with HCI) adjusted to pH 4
(with
NaOH). Four hours after injecting guinea pigs with vehicle (acidified saline
used to dissolve vinpocetine) the animals were anaesthetized and a first set
of
ABR and EEG recordings was taken before the injection (i.p) of the
convulsing agent, PTZ. The animals were injected with PTZ (100 mg/kg) and
about 2 min after injecting PTZ (ictal period) the EEG recording was taken.
Then the other series of ABR and EEG recordings were taken at specific
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times within the post-ictal period. After two weeks the same series of
recordings was taken but instead vehicle the animals were injected with
vinpocetine (2 mg/kg) 4 hours before the injection of PTZ.
The next table shows that vinpocetine inhibits the reduction in P4 wave
peak amplitude induced by PTZ . In the control animals (pre-injected with
vehicle) the amplitude of the P4 wave of the ABR induced by a stimulus of
100 dB at two tone frequencies (8 and 4 kHz) is progressively reduced by
PTZ (left columns), whereas in the animals pre-injected with vinpocetine the
reduction produced induced by PTZ in P4 amplitude is not observed (right
columns). The values shown on the table are the mean standard errors of
the P4 wave amplitudes in pV obtained from 8 animals before and at the
indicated times after the injection of the convulsing agent (PTZ).
8 kHz 8 kHz
PTZ Vinpocetine & PTZ
Before 2.88 0.1 2.69 0.1
10 min 2.07 0.4 * 2.56 0.1
min 1.99 t 0.3 * 2.71+-0.2
min 1.92 0.3 * 2.55 0.2
50 min 1.51 0.5 * 2.56 0.1
4 kHz 4 kHz
PTZ Vinpocetine & PTZ
Before 3.14 0.1 2.82 0.3
10 min 1.70 0.5 * 2.55 0.3
20 min 2.12 0.4 * 2.52 0.2
30 min 1.84 0.4 * 2.55 0.2
50 min 2.11 0.3 * 2.52 0.2
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Vinpocetine also inhibits the increase produced after the injection of
PTZ in the latencies of the ABR waves P2, P3 and P4 induced by a 100 dB
stimulus at 4 and 8 kHz tone frequencies (Fig. 2).
The next table shows that vinpocetine inhibits the hearing loss induced
by the convulsing agent, PTZ. The marked increase on the auditory threshold
at 4 and 8 kHz tone frequencies induced by PTZ (left columns) is not
produced in the animals pre-injected with vinpocetine before PTZ
administration (right columns). The values shown on the table are the mean
standard errors of the thresholds in dB obtained in 8 animals.
PTZ Vinpocetine & PTZ
8 kHz
Before 7.0 1.2 6.0 1.0
30 min 17 2.0 * 6.0 1.0
50 min 21 2.4* 6.0 1.0
4 kHz
Before 21 1.0 20 1.6
30 min 28 2.0 * 18 1.2
50 min 29 1.0* 18 1.2
Vinpocetine inhibits all the changes induced by PTZ on the EEG. All
the anaesthetized animals injected with PTZ developed generalized seizures.
The onset of seizure activity, characterized by repetitive high amplitude
spike-
sharp wave activity in the EEG tracing appears suddenly within the two first
min after PTZ injection. This dramatic change on the cortical activity induced
by PTZ in the anaesthetized animals during convulsions is followed by a
typical pattern of cortical activity characterized by rhythmic spike bursts of
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high amplitude. The duration time of this typical pattern of cortical
activity, that
is not accompanied by convulsions is referred as the post-ictal period.
Vinpocetine completely inhibits the changes on the cortical activity
induced by PTZ for the ictal and post-ictal periods. The top traces on Figs. 3
and 4 show the characteristic EEG recordings under control conditions (i.e.
before the injection of PTZ). The dramatic changes induced about two
minutes after the injection of PTZ on the EEG (ictal period) in the animals
pre-
injected with vehicle are lost when PTZ is injected in the animals pre-
injected
with vinpocetine (Fig. 3). In the same way, the changes induced by PTZ 10,
20, 30 and 50 min after its injection (Fig. 4a) are lost in the animals pre-
injected with vinpocetine (Fig. 4b).
Example 2. Experimental design used for testing the effect of vinpocetine on
the changes on the ABR and EEG recordings induced by an increased
cerebral excitatory transmission.
Five guinea pigs were entered into the study. One hour after injecting
guinea pigs with vehicle (acidified saline used to dissolve vinpocetine) the
animals were anaesthetized and a first set of ABR and EEG recordings was
taken before the injection (i.p) of the convulsing agent, 4-AP. The animals
were injected with 4-AP (2 mg/kg) and about 20 min after injecting 4-AP (ictal
period) the EEG recording was taken. Then the other series of ABR and EEG
recordings were taken at specific times within the post-ictal period. After
two
weeks the same series of recordings were repeated but instead vehicle the
animals were injected with vinpocetine (2 mg/kg) 1 hour before the injection
of
4-AP.
Vinpocetine also inhibits the changes in P3 and P4 waves amplitude
induced by 4-AP. For instance, the progressive increase in the amplitude of
the P3 wave of the ABR produced by 4-AP in control animals (Fig. 5a) is
eliminated in the vinpocetine treated animals (Fig. 5b), and the reduction in
P4
amplitude produced by 4-AP in control animals (Fig. 5c) is markedly reduced
in the vinpocetine treated animals (Fig. 5d).
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The following table shows that the increased latency of the P4 wave of
the ABR induced by the stimulus of 100 dB at 4 and 8 kHz tone frequencies
observed at the indicated times after the injection of 4-AP in control animals
(left columns), is lost when 4-AP is injected in the vinpocetine pre-treated
animals (right columns).
8 kHz 8 kHz
4-AP Vinpocetine & 4-AP
Before 3.48 0.06 3.48 0.04
30 min 3.51 0.13 3.55 0.07
60 min 3.80 0.10 * 3.45 0.08
80 min 3.80 0.10 * 3.36 0.05
100 min 3.83 0.12 * 3.35 0.07
4 kHz 4 kHz
4-AP Vinpocetine & 4-AP
Before 3.50 0.02 3.45 0.06
30 min 3.78 0.09 * 3.37 0.06
60 min 3.78 0.09 * 3.54 0.11
80 min 3.78 0.09* 3.56 0.04
100 min 3.75 0.10* 3.43 0.09
Vinpocetine inhibits the hearing loss induced by 4-AP. The following
table shows that the increase on the auditory threshold induced by 4-AP at 8
and 4 kHz tone frequencies in control animals (left columns) is lost in the
vinpocetine treated animals (right columns).
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4-AP Vinpocetine & 4-AP
8 kHz
Before 3.8 1.3 2.5 1.4
30 min 10.0 2.0 -1.3 2.4
60 min 23.8 6.3 * 0.0 3.5
4 kHz
Before 13.8 1.3 13.8 1.3
30 min 20 2.0 * 11.3 1.3
60 min 30 3.5 * 11.3 1.3
Vinpocetine inhibits all the changes induced by 4-AP on the EEG. All
anaesthetized animals injected with 4-AP developed generalized seizures,
characterized by a repetitive high amplitude spike-sharp wave activity in the
EEG tracing that appears about 20 min after the injection of 4-AP. This
change on the cortical activity elicited by 4-AP for the ictal period is
followed
by the post-ictal period characterized by isolated spikes of higher amplitude
that appear on the EEG about one hour after the injection of 4-AP.
Vinpocetine completely prevents the changes on the cortical activity induced
by 4-AP for the ictal and post-ictal periods. The top traces of Fig. 6 show
the
characteristic EEG recordings taken before the injection of 4-AP in the
animals pre-injected with vehicle (a) and in the animals pre-injected with
vinpocetine (b). The changes induced by 4-AP on for the ictal period are
shown in the second trace on Fig. 6a. In the vinpocetine treated animals 4-AP
was unable to induce the ictal activity (second trace on Fig. 6b). Fig 7a
shows
the post-ictal activity induced by 4-AP, which is lost when the convulsing
agent (4-AP) in injected in the animals pre-treated with vinpocetine (Fig.
7b).
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